SUMMARY At a chemical level, life is inconceivable without metals. This is reflected in the fact that imbalances in metal homeostasis invariably cause disease. Yet, metals are understudied, and their actions are largely taken for granted, in part because the complexity, and systemic integration of metal metabolism and strict cellular reliance on metals pose significant challenges to the exploration of these important contributors to cellular function. Addressing this shortcoming, our research aims to understand the molecular mechanisms that govern the cellular acquisition, distribution and excretion of one particular metal: copper. Although present in only small amounts, the coordination and redox chemistry of copper ions have become indispensible for cells because copper ions enable respiration and detoxification of reactive oxygen species ? two fundamental processes that no aerobically growing organism can live without. Despite much progress in recent years, many fundamental question related to copper metabolism remain unanswered. Through work proposed in this application, we will answer the perplexing question how cells can beat the impossible odds of delivering copper to its final targets, we will develop an in vitro system that reconstitutes cellular copper uptake, and we will make major advances towards determining a structure of the copper pumping Wilson ATPase, ATP7B, that despite intense international efforts has remained an elusive target for structural studies. Aim 1, will be focused on the structure and function of the human copper importer hCTR1. Extending our previous work, we will determine the structure of hCTR1 in complex with CCS, the copper chaperone for Cu,Zn- superoxide dismutase 1, and develop an in vitro assay that will allow us to study mechanistic aspects of copper transport under controlled conditions. Aim 2, will focus on visualizing structure function correlates of the copper pump ATP7B mutations of which are the causative agent in Wilson Disease. Our studies will fill critical gaps in understanding of copper transport across cellular membranes and will advance a new paradigm posing that membrane scaffolding is essential for efficient intracellular copper distribution. Moreover, the anticipated results will make important contributions to understanding the molecular mechanisms underlying disorders that are associated with aberrant copper metabolism such as Wilson's disease, neurodegenerative conditions such as Parkinsons, Alzheimer's and Creutzfeldt- Jakob Disease, and tumor-resistance to platinum-based chemotherapeutics.